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Analysis of hepatitis B virus preS1 variability and prevalence of the rs2296651 polymorphism in a Spanish population

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World Journal of Gastroenterology 2018年第6期24卷 680-692页

作者： Rosario Casillas David Tabernero Josep Gregori irene belmonte Maria Francesca Cortese Carolina González Mar Riveiro-Barciela Rosa Maria López Josep Quer Rafael Esteban Maria Buti Francisco Rodríguez-Frías Liver Pathology Unit Departments of Biochemistry and MicrobiologyHospital Universitari Vall d’HebronUniversitat Autònoma de BarcelonaBarcelona 08035Spain Liver Unit Liver Disease Laboratory-Viral HepatitisVall d’Hebron Institut Recerca-Hospital Universitari Vall d’HebronUniversitat Autònoma de BarcelonaBarcelona 08035Spain Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas(CIBERehd) Instituto de Salud Carlos IIIMadrid 28029Spain Roche Diagnostics SL Sant Cugat del Vallès 08174Spain Liver Unit Department of Internal MedicineHospital Universitari Vall d'HebronUniversitat Autònoma de BarcelonaBarcelona 08035Spain

AIM To determine the variability/conservation of the domain of hepatitis B virus(HBV) pre S1 region that interacts with sodium-taurocholate cotransporting polypeptide(hereafter, NTCP-interacting domain) and the prevalence of the rs2296651 polymorphism(S267 F, NTCP variant) in a Spanish population. METHODS Serum samples from 246 individuals were included and divided into 3 groups: patients with chronic HBV infection(CHB)(n = 41, 73% Caucasians), patients with resolved HBV infection(n = 100, 100% Caucasians) and an HBV-uninfected control group(n = 105, 100% Caucasians). Variability/conservation of the amino acid(aa) sequences of the NTCPinteracting domain,(aa 2-48 in viral genotype D) and a highly conserved pre S1 domain associated with virion morphogenesis(aa 92-103 in viral genotype D) were analyzed by next-generation sequencing and compared in 18 CHB patients with viremia > 4 log IU/mL. The rs2296651 polymorphism was determined in all individuals in all 3 groups using an in-house real-time PCR melting curve *** The HBV pre S1 NTCP-interacting domain showed a high degree of conservation among the examined viral genomes especially between aa 9 and 21(in the genotype D consensus sequence). As compared with the virion morphogenesis domain, the NTCPinteracting domain had a smaller proportion of HBV genotype-unrelated changes comprising > 1% of the quasispecies(25.5% vs 31.8%), but a larger proportion of genotype-associated viral polymorphisms(34% vs 27.3%), according to consensus sequences from Gen Bank patterns of HBV genotypes A to H. Variation/conservation in both domains depended on viral genotype, with genotype C being the most highly conserved and genotype E the most variable(limited finding, only 2 genotype E included). Of note, proline residues were highly conserved in both domains, and serine residues showed changes only to threonine or tyrosine in the virion morphogenesis domain. The rs2296651 polymorphism was not detected in any ***

关键词： Hepatitis B virus Hepatitis B virus preS1 region sodium-taurocholate co-transporting polypeptide NTCP-interacting domain virion morphogenesis domain SNP rs2296651 next-generation sequencing real-time PCR melting curves

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New real-time-PCR method to identify single point mutations in hepatitis C virus

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World Journal of Gastroenterology 2016年第43期22卷 9604-9612页

作者： Qian Chen irene belmonte Maria Buti Leonardo Nieto Damir Garcia-Cehic Josep Gregori Celia Perales Laura Ordeig Meritxell Llorens Maria Eugenia Soria Rafael Esteban Juan Ignacio Esteban Francisco Rodriguez-Frias Josep Quer Liver Unit Internal Medicine Lab. Malalties Hepàtiques Vall d’Hebron Institut Recerca-Hospital Universitari Vall d’ Hebron08035 Barcelona Spain Centro de Investigación Biomédica en Red Enfermedades Hepáticas y Digestivas Instituto de Salud Carlos Ⅲ28029 madrid Spain Liver Pathology Unit Departments of Biochemistry and Microbiology Hospital Universitari Vall d’Hebron Autonomous University of Barcelona 08035 Bareelona Spain Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas del Instituto de Salud Carlos Ⅲ 28029 Madrid Spain Universitat Autònoma de Barcelona 08193 Bareelona Spain

AIM To develop a fast, low-cost diagnostic strategy to identify single point mutations in highly variable genomes such as hepatitis C virus(HCV).METHODS In patients with HCV infection, resistance-associated amino acid substitutions within the viral quasispecies prior to therapy can confer decreased susceptibility to direct-acting antiviral agents and lead to treatment failure and virological relapse. One such naturally occurring mutation is the Q80 K substitution in the HCV-NS3 protease gene, which confers resistance to PI inhibitors, particularly simeprevir. Low-cost, highly sensitive techniques enabling routine detection of these single point mutations would be useful to identify patients at a risk of treatment failure. Light Cycler methods, based on real-time PCR with sequencespecific probe hybridization, have been implemented in most diagnostic laboratories. However, this technique cannot identify single point mutations in highly variable genetic environments, such as the HCV genome. To circumvent this problem, we developed a new method to homogenize all nucleotides present in a region except the point mutation of interest. RESULTS Using nucleotide-specific probes Q, K, and R substitutions at position 80 were clearly identified at a sensitivity of 10%(mutations present at a frequency of at least 10% were detected). The technique was successfully applied to identify the Q80 K substitution in 240 HCV G1 serum samples, with performance comparable to that of direct Sanger sequencing, the current standard procedure for this purpose. The new method was then validated in a Catalonian population of 202 HCV G1-infected individuals. Q80 K was detected in 14.6% of G1 a patients and 0% of G1 b in our setting. CONCLUSION A fast, low-cost diagnostic strategy based on real-time PCR and fluorescence resonance energy transfer probe melting curve analysis has been successfully developed to identify single point mutations in highly variable genomes such as hepatitis C virus. This

关键词： Hepatitis C virus Resistance-associated amino acid substitutions Low-cost test Single-point mutations Q80K

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Neuroprotective effects of meloxicam on transient brain ischemia in rats:the two faces of anti-inflammatory treatments

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Neural Regeneration Research 2023年第9期18卷 1961-1967页

作者： irene Fernández Ugidos Paloma González-Rodríguez María Santos-Galdiano Enrique Font-belmonte Berta Anuncibay-Soto Diego Pérez-Rodríguez JoséManuel Gonzalo-Orden Arsenio Fernández-López Área de Biología Celular Instituto de BiomedicinaCampus de Vegazana s/nUniversidad de LeónLeónSpain Neural Therapies SL.Edif.Institutos de Investigación.Planta baja.Local B43.Campus de Vegazana s/n.León.Spain Department of Medicina Cirugía y Anatomía VeterinariaUniversity of LeónLeónSpain Department of Pharmacology Tulane University School of MedicineNew OrleansLAUSA Department of Neurology David Geffen School of Medicine at University of California(UCLA)Los AngelesCAUSA Department of Life Sciences Imperial College London(ICL)LondonUK Department of Clinical and Movement Neurosciences UCL Queen Square Institute of NeurologyLondonUK

The inflammato ry response plays an important role in neuroprotection and regeneration after ischemic *** use of non-ste roidal anti-inflammatory drugs has been a matter of debate as to whether they have beneficial or detrimental *** this context,the effects of the anti-inflammatory agent meloxicam have been scarcely documented after stro ke,but its ability to inhibit both cyclooxygenase isoforms(1 and 2) could be a promising strategy to modulate postischemic *** study analyzed the effect of meloxicam in a transient focal cerebral ischemia model in rats,measuring its neuroprotective effect after 48 hours and 7 days of reperfusion and the effects of the treatment on the glial scar and regenerative events such as the generation of new progenitors in the subventricular zone and axonal sprouting at the edge of the damaged *** show that meloxicam’s neuroprotective effects remained after 7 days of reperfusion even if its administration was restricted to the two first days after ***,meloxicam treatment modulated glial scar reactivity,which matched with an increase in axonal ***,this treatment decreased the formation of neuronal progenitor *** study discusses the dual role of anti-inflammatory treatments after stro ke and encourages the careful analysis of both the neuroprotective and the regenerative effects in preclinical studies.

关键词： anti-inflammatories astrocyte axonal sprouting cylinder test doublecortin focal brain ischemia glial scar inflammation neuroprotection new neuron generation transient stroke

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